The principal determinant of synaptic serotonin (5-hydroxytryptamine;5-HT) inactivation is the antidepressant-sensitive 5-HT transporter (SERT, SLC6A4). Common polymorphisms in the SERT promoter impact SERT expression and have been associated with anxiety, obsessive- compulsive disorder and autism. SERT is also subject to powerful, posttranslational regulatory mechanisms. Recently, inflammatory cytokines, including IL12 and TNF-1, have been shown to enhance SERT activity in nerve terminal preparations, regulation that can be recapitulated in vivo by systemic cytokine elevation. These findings raise the possibility that SERT may be a convergence point for gene/environment interactions that conspire to alter 5-HT signaling and elevate disease risk. Because SERT is expressed early in brain development, of greatest interest are 5-HT linked disorders of juvenile onset. In this regard, both disrupted 5-HT signaling and disrupted immune function have been advanced as determinants of autism risk. Recently, we identified multiple, functional SERT variants at highly conserved amino acid positions in autism subjects. The most common SERT variant, Gly56Ala, leads to 1) enhanced hSERT catalytic activity, 2) elevated hSERT basal phosphorylation, and 3) a lack of responsiveness to multiple SERT posttranslational regulatory pathways including p38 MAPK pathways activated by inflammatory cytokines. Importantly, the Ala56 hSERT variant was associated with two well-described autism traits, rigid-compulsive behavior and sensory aversion. We have successfully targeted 129S6 mouse embryonic stem cells with a SERT Ala56 "knock-in" construct and obtained germ-line transmission. The SERT Ala56 mice display whole blood hyperserotonemia, hyper-responsiveness to 5HT2A/2C agonist stimulation, and evidence of alterations in social and sensory behaviors. In our current project, we propose a 2-year program to 1) determine the ability of maternal, juvenile and adult SSRIs administration to attenuate the altered behavior and drug responses of 129S6 SERTAla56 mice, 2) to place the 56Ala allele on a C57Bl/6J background using speed-congenic approaches and thereby expand the testable behavioral repertoire of the Ala56 mice, and 3) to establish the sensitivity of SERT Ala56 brain preparations on the C57BL/6J background to in vitro PKG/p38 MAPK stimulation and in vivo stimulation of the native immune system. These studies will enhance our understanding of the contributions of altered SERT activity and regulation to autism heterogeneity and provide a novel platform for the preclinical evaluation of therapeutic interventions. PUBLIC HEALTH RELEVANCE: Rare, functional gene variants that lead to hyperfunction of the serotonin transporter (SERT) have been associated with a specific subset of autism spectrum traits, rigid-compulsive behaviors and sensory aversion. We have produced a novel, transgenic mouse model expressing the most common of these SERT variants, Ala56. Based on exciting preliminary evidence that mice expressing Ala56 SERT exhibit multiple behavioral and biochemical phenotypes, we propose a two-year project to extend our studies and dissect the impact of SERT hyperfunction in vivo.